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From 2009.igem.org

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	LB medium DNA resuspension from iGEM 2009 plates Transformation X-Gal staining protocol for beta galactosidase (blue/white screening) Glycerol stocks Plasmid extraction BioBrick digestion with restriction enzymes Electrophoresis DNA gel extraction DNA precipitation with sodium acetate Ligation PCR Sensing ethanol concentration: our do-it-yourself kit

PCR

(estimated time: 3 hours and 30 min)

For every DNA sample you want to amplify, put: 2 µl buffer 0.6 µl MgCl2 0.4 µl dNTPs 1 µl DNA (or ddH2O for blank sample). If you are performing a colony PCR, pick up the desired colony from a plate with a tip and dip it in the solution. 0.2 µl Taq Polymerase 250 nM VF2 primer 250 nM VR primer A proper amount of ddH2O to have 20 µl of total reaction volume into an eppendorf tube. Put the eppendorf tube in the thermal cycler and set this program: 95°C 10 min CYCLE: 95°C 30 sec 60°C 1 min 72°C 1-3 min for 35 cycles 72°C 7 min 16°C forever. Now you can add a loading buffer to the solution and perform electrophoresis to check the amplified sequence length.

MgCl2 Buffer dNTPs ddH2O Taq Polymerase VF2 primer VR primer

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